Hang Yao, Yuwei He, Jinrong Ma, Lang Jiang, Jingan Li, Jin Wang,*, Nan Huang

1 Key Laboratory of Advanced Technology for Materials of the Education Ministry, School of Materials Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China

2 Department of Chemical Engineering, University of Washington, Seattle 98195, USA

3 Department of Pharmaceutics, School of Pharmacy, Fudan University & Key Laboratory of Smart Drug Delivery, Ministry of Education, Shanghai 201203, China

4 School of Materials Science and Engineering, Zhengzhou University, Zhengzhou 450000, China

Keywords:Cardiovascular stent modification In-stent restenosis Late stent thrombosis Re-endothelialization Inflammatory modulation

ABSTRACT Treatments of atherogenesis, one of the most common cardiovascular diseases (CVD), are continuously being made thanks to innovation and an increasingly in-depth knowledge of percutaneous transluminal coronary angioplasty (PTCA), the most revolutionary medical procedure used for vascular restoration.Combined with an expanding balloon,vascular stents used at stricture sites enable the long-time restoration of vascular permeability.However,complication after stenting,in-stent restenosis(ISR),hinders the advancement of vascular stents and are associated with high medical costs for patients for decades years.Thus, the development of a high biocompatibility stent with improved safety and efficiency is urgently needed.This review provides an overview of current advances and potential technologies for the modification of stents for better treatment and prevention of ISR.In particular, the mechanisms of in-stent restenosis are investigated and summarized with the aim to comprehensively understanding the pathogenesis of stent complications.Then,according to different therapeutic functions,the current stent modification strategies are reviewed, including polymeric drug eluting stents, biological friendly stents, prohealing stents,and gene stents.Finally,the review provides an outlook of the challenges in the design of stents with optimal properties.Therefore, this review is a valuable and practical guideline for the development of cardiovascular stents.

1.Introduction

Percutaneous transluminal coronary angioplasty (PTCA) is a minimally invasive procedure designed to treat narrowing vascular vessels with atherosclerosis via balloon angioplasty.Sigwart et al.first introduced metallic stents to remold vessels by providing them with long-term support in 1987 [1].Although PTCA with stent implantation represents a technological breakthrough, leading to the replacement of the coronary artery bypass graft surgery,it is associated with various serious complications, including instent restenosis (ISR), late stent thrombosis (LST), and bleeding after expensive dual anti-platelet therapy (DAPT) for more than 1 year [2].In addition, due to the rejection of stents via immune response, stent materials have become a driving factor of these complications.Additionally, the elastic expansion of stents can result in continuous physical damage to vessels and the subsequent series of injury-induced healing responses.

To date, many researchers studying surface modification have focused on the design of novel cardiovascular stents in compliance with the complex in vivo environment.However, no one has been able to eliminate the previous issues completely without creating new problems.For example, the first successful cardiovascular stent implantation is bare mental stent (BMS) intervention, made using 316L SS, effectively overcome artery elastic recoil, acute thrombus, and myocardial infarction post-coronary angioplasty.However, the clinical incidence of ISR due to excessive neointimal hyperplasia was about 30%after 2-3 months BMS deployment[3].Then, the first-generation drug eluting stent (1G DES) was introduced, delivering drugs towards the hyperplasia site to inhibit excess proliferation.However, complications after DAPT suspension occurred in about 3 % of patients who underwent DES within 1 year, raising serious questions with regards to the safety of 1G DES [4].In recent decades, numerous researchers have focused on modifying DES to improve its safety profile.Unfortunately,although new generation DESs (second generation (2G) and third generation (3G)) show better long-term safety profiles, thanks to the replacement of biostable polymer coatings with biocompatible or bioresorbable polymers, the application of high elasticity modulus metal to produce thinner stent struts,and the use of less cytotoxic drug(e.g.everolimus(vs.sirolimus&paclitaxel)),the LST rate for DES is still higher than BMS,the gold standard of safety.In general, LST is closely associated with delayed endothelial layer healing, local innate inflammation, the toxicity of anti-proliferation drug, and the physical damage caused by stents to surrounding vascular tissues [5].Furthermore, bioresorbable vascular scaffolds(BVS) (Abbott Vascular), which are the most widely used bioresorbable stent (BRS), have a higher scaffold LST rate (0.2 % to 1%) after 24 months compared to new generation DESs [6].

Thus, neither DES nor BRS resolve ISR satisfactorily without causing other issues.Now, concerns over ISR have resurfaced,accompanied by the belief that a better approach must exist for the treatment of ISR, and not simply through the delivery of antiproliferation drugs.Therefore, ISR remains an essential problem,even in the era of cardiovascular stents.In order to design a cardiovascular stent with a higher safety and efficiency, potential therapeutic targets that can effectively suppress neointimal hyperplasia while minimizing adverse complications should be investigated.To this end,this review summarizes various currently used strategies and potential therapies,as shown in Fig.1,with the aim of paving the way for a solution to ISR without giving rise to other complications.

2.Mechanism of In-Stent Restenosis

Fig.1.The overview of structures in this review.According to the different therapeutic targets, the current stents are reviewed including drug eluting stent,biologically friendly stent, pro-healing stent, and gene therapy as shown in the inner circle and modification strategies are summarized as shown in the external circle.

ISR involves an artery becoming miss-shaped and hardening due to interactions between inflammatory cells,platelets,extracellular matrix (ECM), smooth muscle cells (SMCs), and endothelial cells (ECs), resulting in the re-blocking of the artery [7-9].There are three stages of ISR development, as shown in the Fig.2: (1)early stage (≤2 months): due to the vascular injury, mild thrombi deposit on the material’s surface, followed by the accumulation of inflammatory cells,coagulation,the formation of early thrombosis,and acute inflammation;(2) mid stage (2-18 months): SMCs start to rapidly proliferate, evolving into progressive neointimal hyperplasia; (3) late stage (≥18 months): lastly, neointima stabilize and regress [10,11].Conventional IRS therapies mainly focus on the suppression of SMC proliferation and migration, such as DES.However.ISR is a complicated pathological process with multifactorial triggers.Herein, we provide an integrated insight into the mechanisms underlying the development of ISR.

2.1.Smooth muscle cells and extracellular matrix

It is well known that neointimal tissues are mainly comprised of SMCs and ECM.As shown in Fig.2, stent struts are covered by a thin,membranous thrombus after implantation in vascular vessels.Then, SMCs increasingly proliferate, secreting ECM to the surrounding tissues.The dominant component of neointima is alpha-actin-positive SMCs at the early stage[12].In the late stage,the replication of SMCs tends to terminate, transforming into the stable phase.After cell death, the depleted area increases and the ECM starts to accumulate, filling in any empty areas [13].Chung et al.found that the extrusion and compression of vascular tissues when stents expanded excessively was a contributor to ECM remolding [14].Changes in the matrix protein components may play an important role in ECM remolding.Upon the stimulation of TGF-β1, the increased synthesis of proteins such as proteoglycan, elastin, and collagen Ⅰresults in the disruption of matrix homeostasis, resulting in ECM remodeling [15-17].Recently, Suna et al.demonstrated a that aggrecan in ISR was significantly upregulated by proteomics technology, and concluded that aggrecanase activity plays an important role in vascular injury responses poststenting,which may provide a novel drug target for the regulation of ECM remolding [7].Thus, the excessive proliferation of SMCs contributes to early stage ISR,while ECM remolding is an essential factor leading to late stage ISR.

2.2.Inflammatory response

Innate inflammation occurs through the ISR process, reaching its maximum level at late stage [18].As shown in Fig.2, platelets and fibrin are directly deposited onto the surface of stents poststent deployment, followed by the formation of a thin membranous thrombus.Inflammatory cells (i.e.leukocytes and macrophages) are then recruited by P-selectin and glycoprotein Ibα from active platelets,as well as fibrin,and adhere to platelet-fibrin networks, resulting in acute inflammation [18,19].Growth factors(i.e.VEGF, PDGF, IGF, and TGF-β) are then secreted from activated inflammatory cells, after which platelets and SMCs further simulate the neointimal hyperplasia [20,21].Although hyperplasia intima can act as barrier to isolate inflammatory cells, driven by a chemical chemokine gradient (i.e.interleukin (IL)-8), cells can still infiltrate via the neointima slowly, gradually evolving into chronic inflammation.At the late stage, the inflammatory responses are enhanced by complex matrix components.

2.3.Endothelium cells

Fig.2.Illustration to vascular responses after stent implantation in time-sequence order from left to right.At early stage(≤2 months),the vascular vessel is inevitable injured after stent deployment and the exposure of media layer triggers acute thrombosis by activating platelets and fibrinogen adhesion;At mid stage (2-18 months),the smooth muscle cells(SMCs)start maniacal proliferation around stent surface which induce obvious re-stenosis in vessel;At late stage(≥18 months),Since enormous inflammatory cells infiltration(i.e.macrophage,neutrophil and foam cell),the inflammation intensively appears at this stage and extracellular matrix(ECM)is remolded by secreting to fill the spare room of neointima.

After the implantation of stents, excess expansion can lead them to penetrate vessel walls, resulting in severe and constant damage to the endothelial layer.The momentary elastic expansion of stents in vessels can destroy endothelial vessel walls completely.With regards to long-term stents, the phase of endothelial layer denudation can last up to 12 weeks, followed by unintegrated reendothelialization on parts of the neointima.The degree of neointimal hyperplasia is proportional to the area of denudation.Larger denudation areas are indicative of serious neointimal hyperplasia [22].Compared with balloon angioplasty, delayed endothelial layer healing is observed in stent implantation, featuring dysfunctional ECs and leukocyte adhesion [23,24].Although ECs are completely covered at the late stage of ISR, the absence of endothelium at the early-and mid-stage may be an important contributing factor to ISR,since mature and healthy ECs serve not only as a physiological barrier against ECM accumulation but also as regulators of SMC proliferation by secreting nitric oxide (NO)[25,26].Additionally, endothelial shear stress (ESS), the tangential stress derived from the friction between flowing blood and the endothelial surface, is another potential contributor to ISR[27,28].Stents alter local artery geometry and EES patterns concomitantly, as shown in Fig.3, leading to low EES at the downstream side of the strut, especially at the reign of proximal and distal sites[29-31].Jimenez et al.reported that a streamlined stent strut design would promote the development of a local flow field free of recirculation zones, thereby avoiding the appearance of low EES in vessels [32].Due to the difficulty of measuring EES in arteries, imaging diagnostic tools are expected to promote advances in this field in clinical settings.

3.Drug-Eluting Stents

Patients with BMS usually develop severe restenosis 2-3 months after stent implantation.Up to 25 % of patients who received metal stents developed ISR post-stent implantation.Drug eluting stent(DES)was found to significantly reduce the risk of ISR by delivering anti-proliferative drugs to inhibit the abnormal proliferation of SMCs, a vital factor in the development of ISR.In traditional DES systems, stents are composed of three parts: (1)metallic platform;(2)polymer coating;(3)anti-proliferative drugs.The metallic platform undergoes plastic deformation and should maintain the size required at the target site.Thus, the metallic platform should meet several requirements, including a low profile, good expandability ratio, flexibility, and sufficient radial strength [33].Conventional stent platforms are generally made of stainless steel or tantalum alloy.Alternative alloys, such as cobalt-chrome (Co-Cr), nitinol (Ni), and platinum-chrome (Pt-Cr), have also been used to improve radial strength with thinner struts that favor reduced vascular injury post-stent expansion and the disruption of blood flow [34,35].Clinical studies have shown that thinner struts are conductive to ISR prevention, such as Multi-link Vision Co-Cr (L605) platform (Abbot Vascular, CA,USA) and Driver Co-Ni-Cr (MP35N) platform (Medtronic, CA,USA) [36,37].Thus, BMS and metallic platforms should use strong metal materials for the production of ultrathin struts.In this review, since polymeric coating is a very important component of DES systems (i.e.necessary to achieve drug adherence on the metallic surface and to control drug release kinetics),we introduce traditional DESs in-depth according to the types of drug-carrier polymers.

Fig.3.Overview of micro flow environment around a stent strut (left) and the resulting high and low wall shear (right).

Fig.4.FDA approved 1G & 2G drug eluting stents.

3.1.Biostable polymeric drug-eluting stents

For DESs as shown in Fig.4,both 1G and 2G DESs belong to biostable polymeric DESs and have been approved by U.S.Food and Drug Administration (FDA).For 1G DES, the most representative stents include Taxus stent (Boston Scientific, Natick, MA, USA)[38]with poly(styrene-b-isobutylene-b-styrene) (SIBS) copolymer loading paclitaxel and Cypher Stent (Johnson & Johnson, New Brunswick, NJ, USA) [39]with a blend of two durable polymers(poly (ethyleneco-vinyl acetate) (PEVA) and poly(n-butyl methacrylate)(PBMA))to load sirolimus.These two stents are both based on stainless steel platforms[40,41].Taxus was the first FDA approved DES used in stent market and was expected to replace BMS.Although 1G DES achieved great commercial success, reducing the rate of ISR below 5 %[42],significant deficiencies emerged after extensive clinical implantations, including uncontrolled drug release, early (＜4 weeks), late (≥1 year), and very late (≥2 year)thrombosis due to their inferior hemocompatibility, and delayed endothelial layer healing [40,43,44].The design of 1G DES places a greater emphasis on the compatibility between the drug and polymer matrix, the duration of elution, and the release kinetics.By substituting stainless steel with Co-Cr or Pt-Co alloy with a thinner strut thickness, as well as an improved strength and radio-opacity, 2G DES allows for a lower risk of restenosis in clinical settings.2G DES also represents an upgrade on the polymeric coatings, with more biocompatible polymer that are still durable(e.g.PC and PVDF-HFP),allowing for a longer duration of drug elution and steady release behaviors.Furthermore, drugs with stronger anti-proliferation effects, such as everolimus and tacrolimus,can be applied to the eluting system.These improvements have led to 2G DES replacing 1G DESs in clinical use.Xience V Everolimus-eluting stent (Abbott Laboratories, Abbott Park, IL,USA)is one of the most prominent 2G DES(Fig.4),and is regarded as the golden standard of coronary stents due to its excellent performance in clinical applications [45,46].Table 1 provides a list of all current FDA approved 1G and 2G DESs [47-50].Although 2G DES has significantly reduced the LST risk compared to 1G DES[51,52],the risk of very late stent thrombosis(VLST)remains after 2 years in patients with the new generation DES.Although the precise reasons for VLST are not yet fully understood, polymerinduced inflammation and delayed endothelium repair in vivo are thought to be responsible [18,53].Given the evidence of the long-term safety issues induced by durable DES polymers, strategies to replace biostable polymeric carriers with biodegradable polymers and biological-friendly polymers are discussed in the following sections.

3.2.Biodegradable polymeric drug-eluting stents

To address the challenges associated with biostable polymer coatings,including inflammation,ISR,and LST,biodegradable polymer(BP)coatings are used in BP-DES,allowing for a control release matrix for drug elution at the early and medium stages, which degrade at the late stage of implantation.The main commercial biodegradable polymers used for DES coatings are polylactic acid(PLA), poly (l-lactic acid) (PLLA), and copolymer polylactic-coglycolic acid(PLGA).For example,Biomatrix stent(Biosensors Europe, Morges, Switzerland) was designed to elute Biolimus A9, an anti-proliferation drug with a therapeutic efficiency stronger than sirolimus, from PLA coating matrices.Table 2 summarizes current CE approved the third generation(3G)DESs with degradable polymeric coatings[54-59].This approach has been shown to improve long-term safety and efficacy compared to 1G DESs, but have not been shown to have a performance superior to 2G DESs[60].Thus,it is too early to quantify the success of the new generation of DESs.One three-year follow-up clinical study for the left main coronary artery did not find any significant differences between 2G DESs(CoCr-Everolimus DES, PtCr-Zitarolimus DES, Re- Zitarolimus DES) and 3G DESs (Biomatrix stent) in terms of the risk of target vascular failure and other adverse cardiac events [61,62].More recently,a 1-year clinical report found that biodegradable polymer sirolimus eluting stents were superior to durable polymer everolimus eluting stents in terms of target lesion failure in patients with acute STEMI [63].

Table 1 Summary of the first and second generation drug eluting stents

Table 2 Summary of the third generation drug eluting stents

Table 3 Summary of current vectors used to treat cardiovascular diseases

3.3.Bioactive polymeric drug-eluting stents

In traditional thrombogenic 1G and 2G DESs, proteins (fibrinogen, von Willebrand factor, albumin, and fibronectin) rapidly adhere onto the surface of stents upon contact with the vascular blood.The non-specific adhesion of proteins can trigger a series of adverse events, including platelet adhesion, activation, inflammation, excess proliferation, and thrombosis.Thus, suppressing protein fouling is an attractive strategy to reduce polymer-biological environment interaction and prevent the formation of thrombogenic biofilms [64,65].DAPT is a prominent and effective clinical therapy resistant to thrombosis post-PTCA surgery.However, patients undergoing DAPT are required to take anti-platelet drugs (e.g.asplin, clopidogrel, or ADP receptor antagonist) continuously for up to two years,at a huge economic cost.Therefore,the development of a bioactive polymeric coating for DES is needed to improve stent biocompatibility and the cost-effectiveness of treatment.Anti-fouling polymers are an attractive alternative for a bioactive polymeric coating, providing resistance against the non-specific adsorption of proteins and thrombogenic biofilm formation by reducing the non-specific interactions between materials and the biological environment.Most biological anti-fouling polymers currently used in cardiovascular stents are derived from natural substances and have good biocompatibilities, including phosphorylcholine (PC), hyaluronic acid (HA), and fibrin matrix.In addition to those derived from natural materials,many synthetical materials also exhibit excellent anti-fouling properties,such as polyethylene glycol (PEG), poly-sulfobetaine (PSB), and polycarboxybetaine (PCB).Given their outstanding biocompatibility with reduced foreign reactions, they are widely used as potential drug or biomolecule delivery platforms [66].

Zwitterionic polymer is an electroneutral material polymerized by zwitterion monomers with a functional group containing one positive electrical charge and one negative electrical charge.By forming strong electrostatically-induced hydration on the surface of materials, zwitterionic polymers maintain the conformation of biomacromolecules and resist the adsorption of non-specific proteins[67,68].Thus,zwitterionic polymers are a promising alternative blood contact material to reduce interactions between polymers and the biological environment and the formation of thrombogenic biofilm [69,70].2-methacryloyloxyethyl phosphorylcholine (MPC) is one of the most popular zwitterions used in medical devices [71].Poly-MPC has a good biocompatibility and hemocompatibility for use in cardiovascular stents.The Biodiv Ysio stent(Abbott Laboratories,Abbott Park,IL)has excellent long-term antithrombogenic properties (≥6 months), utilizing PC as a stent coating regardless of the drug loaded [72].Thus, PC is considered as an attractive drug delivery platform for DES due to its safety and biocompatibility on the long-term, as well as it anti-coagulation properties.Endeavor DES (Medtronic Inc., Minneapolis, MN, USA)was the first 2G DES(approved by FDA in 2008)with a PC coating,a CoCr metal platform, and zotarolimus drug (Fig.4).A four-year clinical follow-up trial found that Endeavor-ZES exhibited continuously low rates of major adverse cardiovascular events (MACE),target lesion recurrence(TLR)[73],and noninferior efficiency compared to Paclitaxel eluting stent(PES)[74].However,another clinical study found that Endeavor-ZES had a less early efficiency compared to everolimus eluting stents,but more regression of yellow plaque, which play a key role in the development of neoatherosclerosis, indicating that Endeavor-ZES has a better late safety profile[75].In a preclinical study,hyperbranched polymeric coating combined with 2-((bromobutyryl)oxy)ethyl acrylate(BBEA) and the zwitterions 3-dimethyl(methacryloyloxyethyl)ammonium propane sulfonate (DMAPS) was reported by Wang et al., as shown in Fig.5A.This results in improved blood compatibility via atom transfer radical polymerization (ATRP) [76].The Ji group fabricated an multifunctional coating by synthesizing a segmented copolymer containing MPC, n-stearyl methacrylateand,and p-nitrophenyloxycarbonyl poly(ethylene glycol) methacrylate units.The REDV peptide was conjugated onto copolymer surfaces to selectively promote endothelial layer healing.Meanwhile, the PC unit was able to effectively resist protein fouling to improve hemocompatibility [77].Furthermore, The Ji group reported a novel biodegradable MDO-PC-BMA copolymer via one-step radical ring-opening polymerization, which showed excellent biocompatibility, biodegradability, and rapamycin-eluting capacity [78].

In addition to zwitterionic polymers, hyaluronic acid (HA), a non-sulfated glycosaminoglycan distributed widely throughout the ECM of soft connective, fluid, and tissue, is recognized as another top candidate for use in anti-fouling coatings [79,80].Verheye et al.first quantified the biological effects of HA for use as a potential stent coating under physiological blood flow conditions.They concluded that HA-coated devices significantly reduced platelet adhesion and aggregation ((0.82 ± 0.20) × 109VS (1.83 ± 0.23) × 109platelets; P ＜0.02), confirming that HA is an attractive drug carrier matrix for improving the thromboresistance of stents[81].In addition to its anti-thrombosis properties,HA was reported to decrease the inflammatory responses post-stent implantation,most likely due to its antioxidant properties [82].Interestingly,HA coatings were reported to synergistically enhance the modulation ability of other biological agents, such as type IV collagen, by enhancing the modulation of type IV to the contractile phenotype of SMCs by formulizing the collagen order[83].As shown in Fig.5B,Yu et al.established a multifunctional dopamine modified surface wherein the amino group covalently bonded with the aldehyde group of MA-HA.Then, a high-density functional Arg-Glu-Asp-Val peptide (REDV) was conjugated via thiol-ene to selectively promote EC adhesion and growth.Compared with polystyrene, the multifunctional coating was confirmed to resist the adhesion of SMCs up to 25 % (P ＜0.005) that of polystyrene and promote EC migration [84].These results highlight the great commercial potential of bioactive polymers for improving the biocompatibility of polymeric stent coatings.

4.Biologically Friendly Stents

Vessels are usually concomitant with acute and chronic inflammatory reactions post-stent implantation.This plays a pivotal role in the development of ISR,as illustrated above.Inflammatory cells,such as macrophages and lymphocytes, accumulate in large amounts around stents at both the early and late stage after stenting[85,86].Macrophages,lymphocytes,and SMCs secrete vascular endothelial growth factor-A(VEGF-A)and platelet-derived growth factor-BB(PDGF-BB)to accelerate SMC proliferation[21].Thus,the development of biologically friendly stents is a promising strategy to reduce the amount of inflammation associated with cardiovascular stents.

4.1.Fully bioresorbable stents

Although 1G and 2G DESs are able to reduce the incidence of ISR below 5 % via drug delivery, polymers can trigger inflammatory responses, and are closely linked with an increased risk of LST[87,88].Studies have found that polymer-induced inflammation is correlated with poor polymer biocompatibility, polymer degradation products,including acids,initiators,and catalysts,and coating defects, such as cracking by stent expansion [89].Completely bioresorbable stents are designed to be biofriendly to the surrounding microenvironment.BVS 2.0(Abbott Vascular,Santa Clara,CA, USA) was approved by FDA in 2014 and is one of the most famous disappearing stents.It is comprised of biodegradable PDLLA polymeric stent bone and everolimus loaded to minimize stimuli to the vascular tissue.It is expected to maintain adequate mechanical radical power and control drug release at the early and mid stages,and fully disappear in 6 months,preventing physical damage to vessels and polymer-induced inflammation.Although it is a promising and revolutionary stent and has obtained good clinical results before making his debut in market,the increased incidence of LST after many implantations is likely to hinder the broad application of BVS, among others [90-92]: (1)the thrombogenicity of PDLLA polymeric matrix;(2)the high physical profile of stent increases the operating difficulty for doctors;(3) disturbed blood flow due to the lack of complete integration after degradation; (4) prolonged inflammatory responses detected due to slow degradation rate.

Fig.5.(A) Schematic illustration to show the surface-initiated ATRP of DMAPS from bare metal stent (BMS) surface to produce the BMS-g-HPBBEA-b-PDMAPS modified surface.(B)Schematic illustration to show the EC-selective peptide(REDV)density gradient created on a uniform methacrylate anhydride-modified hyaluronic acid(MA-HA)layer, which is constructed on top of a polydopamine-coated poly(ε-caprolactone) (PCL) film.

Biodegradable metals (i.e.Mg, Fe, and Zn) have been proposed due to higher mechanical properties than bioresorbable polymers and non-toxic degradation products, without triggering a marked inflammatory response.However, the slow degradation rate of iron-based alloys and the rapid un-uniform degradation of magnesium alloy (0.1-0.2 mm·a-1) are some of the limitations of biodegradable metals applied to cardiovascular stents [93].Wang et al.established a polymeric coating on a magnesium alloy surface to reduce the degradation rate [94].The corrosion current density of the poly(1,3-trimethylene carbonate) (PTMC)-coated alloy was reduced by three and one order of magnitude compared to bare and poly(e-caprolactone) (PCL)-coated magnesium alloy, respectively.A homogeneous surface without pores or cracks was observed, as shown in Fig.6A.Erosion occurred from the exterior to the interior of alloy due to the protective function of the PTMC coating.Sheng grafted a zwitterionic phosphorylcholine chitosan(PCCs)polymer to the surface of magnesium alloy, endowing the materials with an improved biocompatibility and anti-corrosion behavior simultaneously [95].Changing the composition of alloys is used to improve the resistance of stents to corrosion.Lin et al.reported a promising composition of magnesium alloy [96].The JDBM-2 stent, comprised of Mg (2.2%, mass), Nd (0.1%, mass), Zn(0.4%,mass),and Zr(showed a low toxicity,slow erosion rate with up to 6-month structural,mechanical integrity, and homogeneous erosion compared to conventional AZ31 magnesium alloy(Mg(3%,mass), Al (1%, mass) Zn, and Mn (0.2%, mass)).Furthermore, zincbased alloys have been studied in recent years as a promising biodegradable metal.Because of the tolerable corrosion rate(0.02 mm·a-1) and better biocompatibility of zinc, zinc-based metallic stents have been the focus of extensive research [97,98].Bowen et al.found that pure zinc wire did not induce a negative inflammatory response or SMC proliferation up to 6 months after implantation in rats in vivo [99].However, the high elongation to failure of pure zinc increases its processing difficulties [100,101].Ma et al.studied the cellular responses of human coronary artery endothelial cells (HCECs) under the influence of Zn2+up to 0.14 mmol·L-1, and found that low Zn2+concentrations benefitted the proliferation, adhesion, and migration of HCECs [102].Moreover, various alloy, such as Zn-Cu, Zn-Al, and Zn-Ca, have been extensively investigated with regards to their influence on corrosion behavior,with the aim of finding a corrosion rate that matches the tissue healing rate [103].

Fig.6.(A) The schematic of erosion behaviors of PTMC coated magnesium alloy (left) and bare magnesium alloy metal (right).(B) The scheme of photoactive super hydrophilic nanofilm with NO generation and self-cleaning properties.

4.2.Polymer-free stents

Another route for establishing biologically friendly stents involves removing the pro-inflammatory stimuli from the stent system.In other words, the development of a polymer-free stent.To attain the equivalent drug release behavior as polymeric DESs,the drug coating methods are a great challenge for polymer-free stents.Crystallization is an effective way to control drug release behavior via micro-drop spray crystallization or temperatureinduced crystallization [104].The crystal structure improves the burst release control at the early stage since a stable crystal enables a slower dissolution rate than that of the amorphous phase.However, the crystallization approach remains unable to achieve a long-term controlled release profile.Thus, the physicochemical properties of drugs need be considered when establishing a stable crystal phase.Furthermore, building macro-, micro-, and nano-pore reservoirs or grooves on the surface of the stents is another effective alternative.This allows for larger dosages,as well as enhancing the adhesive strength between the drug and the metallic platform [105,106].Additionally, the diffusion distance is increased,while the exposure area of drugs localized to abluminal grooves is decreased.All of these methods are conductive to an elevated controlled release profile in polymer-free stents [107].Recently, a promising clinic study revealed that polymer-free biolimus eluting stent(PF-BES)showed a similar safety and efficiency profile as Xience CoCr-Everolimus eluting stent [108].As the world’s lowest drug dosage classic workhorse stent, with a dosage that is 1/3 of peer products, the Firehawk Rapamycin Target Eluting Coronary Stent System (MicroPort Medical Inc.Shanghai,China)achieves a similar efficacy in terms of the primary endpoint of ISR after 13 months compared to XIENCE durable polymeric everolimus eluting stents [109].The abluminal grooves contain bioresorbable PDLLA polymer acting as a carrier for sirolimus.The biodegradable polymeric matrix exposed area is only 5 % of the stent surface, ensuring the controlled released of rapamycin over 90 d and its full degradation within 9 months in vivo.Since most of the polymer is comprised of abluminal grooves and only a small area is exposed to the abluminal vessel, the influence of polymer is minimized compared to fully-covered DES systems.Thus, an ultra-low drug dose and polymer exposure prevents amplifying inflammation and the toxicity of drugs.

4.3.Biomimetic stents

Biomimicry of the endothelial layer is a powerful strategy for regulating vascular responses after stenting.Nitric oxide (NO)derived from ECs has become a popular biomimetic target due to its versatile functions.To date,NO gas has been found to suppress the excess growth of SMC, resist platelet activation via cyclic guanylate monophosphate(cGMP)pathway,and modulate inflammatory reactions [110,111].To make use of healthy endothelial layer functions for vascular healing post-stent deployment, in situ NO-generation materials were constructed by inhaling NO gas or loading metallic catalyst(e.g.Cu and Se)to decompose endogenous NO donors in the vascular blood [112-114].Kushwaha et al.utilized a self-assemble peptide amphiphile as the matrix to inhale NO molecules.This endothelial ECM mimicked self-assembled nanofibrous coating and successfully inhibited platelet activation,adhesion,and SMC proliferation and delayed reendothelialization[115].The Huang group developed a biomimetic coating on cardiovascular stents with synergistic effects by combining heparin and selenocystamine catalyzed NO gas [116].NO generation coupled with heparin synergistically enhanced the anti-thrombosis ability, and suppressed SMC proliferation by simultaneously upregulating α-smooth muscle actin expression and cGMP synthesis.Recently, a copper-mediated photoactive anti-protein fouling TiO2nanofilm was proposed by the Huang group that enabled well-retained NO catalytic sites to improve the NO release efficiency in vivo,as shown in Fig.6B.The enhanced modulation of inflammation by the synergistic effects of NO generation and anti-protein fouling were highlighted in this work[117].

5.Pro-Healing Stents

Delayed endothelial healing after stenting is closely associated with ISR.From a pathophysiological perspective, normal ECs can maintain vessel wall homeostasis by producing vasoactive NO,prostacyclin (PGI2), and other modulatory agents to protect the vessel wall against inflammatory cells and platelets adhesion,thrombus formation, and SMCs proliferation [118].However, the inevitable and long-term damage of vessels during stent service involves the endothelial layer underdoing a prolonged healing process, wherein EC dysfunction can occur [23].Growth inhibiting mediators are usually absent from dysfunctional ECs, leading to the proliferation and migration of SMCs in the intimal space.NO is markedly decreased,while cell surface adhesion molecules,such as selectins and integrins, are intensively synthesized, increasing the inflammatory response and adhesion of platelets to stents.Importantly, Kipshidze et al.has confirmed that delayed endothelial healing due to EC damage is a crucial reason for neointima formation in-stent.In turn, rapid re-endothelialization has potential for inhibiting ISR without the development of late thrombus[119,120].Herein, we review several rapid re-endothelialization strategies for stent modification.

5.1.Surface micro- and nano-pattern stents

Surface topography in microscale or nanoscale ranges can be used to immobilize cells onto the surface of stents.Once seeded,cells can migrate to the top of the ridges rather than the top of grooves (contact guidance) [23].Several studies have suggested that the specific microstructure of surfaces promotes cell adhesion and proliferation, which are connected with cellular contraction, stress fibers, and focal adhesion [121,122].Compared with SMCs, ECs are known to prefer microstructure patterns.Femtosecond Laser (FSL) technology can be used to produce precise microstructures on the surfaces of metallic stents via light pulses of extremely short duration.The use of FSL with a 3D rotating system was reported to improve re-endothelialization compared to commercial gold standard stents (everolimuseluting stent) [123].As shown in Fig.7A, Mingdong et al.designed a special SMC-biomimetic microstructure pattern using FSL on a 316L SS stent for EC migration and growth.The fabricated SMC pattern (LTS) with a fiber width of ～700 nm is able to selectively promote the adhesion, proliferation, and migration of ECs, and is indicative of a promising surface pattern with endothelium pro-healing functions [124].Additionally, chemical vapor deposition refers to an effective method to produce patterns on the surface of polymeric matrices.A newly designed surface pattern MPC polymer was reported by Kenta Bito et al.,who produced a micropatterned hydrogenated amorphous carbon (a-C:H) pattern via chemical vapor deposition, as shown in Fig.7B (left) [125].The micropatterned a-C:H/MPC polymer matrix effectively supported the proliferation of ECs and regulated the migration of ECs by changing the size of the C:H island,as shown in Fig.7B (right).

Fig.7.(A)Comparison of the EC on different surfaces(a)SEM image of the cells growing on NMS and LMS;(b,c)Zoomed-in images from the dashed white squares in(a);(d,e) The fluorescent images of the EC on NMS and LMS; (B) The schematic image showing the process of patterned MPC polymer with different sizes and behaviors of EC cultured with the micropatterning polymer.

5.2.Dual drug-eluting stents

Based on the DES system, eluting re-endothelialization agents can be used to promote re-endothelialization.A widely used biomolecule is the vascular endothelial growth factor (VEGF), which promotes EC growth and reduces thrombus risk [126,127].Additionally, several novel drugs, such as estradiol, arsenic trioxide,and vildagliptin, may exert re-endothelization functions in DESs[128-130].However, the strategy of solely delivering reendothelium agents is typically insufficient to inhibit the proliferation of SMCs.For example, a VEGF eluting stent was reported to wrong target SMCs to promote undesired hyperplasia.Although DESs loaded with anti-proliferative drugs may reduce early ISR,the high risk of LST is the main limitation of 1G-DES and 2G-DES.Anti-proliferative drug damage ECs unselectively while inhibiting SMCs proliferation, leading to drug-induced deficient reendothelialization [131-133].Dual drug-eluting stents (DDES)have been developed to tackle this problem by taking advantages of both anti-proliferation and pro-healing drugs, such as Paclitaxel-VEGF [134].Using the synergistic effects of two bioactive agents incorporated in a polymeric coating or polymer-free reservoir,clinical studies have reported their potential in overcoming very late thrombosis without comprising the anti-ISR efficiency, compared to 1G DESs, such as sirolimus-probucol,sirolimus-triflusal, and atorvastatin-fenofibrat DDESs [135-138].The ideal drug kinetics of DDES involves sequential eluting behaviors, wherein the stent elutes two drugs separately at different stages,with the aim of matching the eluting kinetics of the vascular healing process.A pioneering DDES strategy in which drugs were eluted separately was reported by Du et al., who fabricated novel SZ-21/DTX drug-loaded hydrophobic core/hydrophilic shell particle coating stents via electrospraying.Therein, the antiproliferative drug docetaxel was located in the core,while the antibody SZ-21 was located in the shell [139].This sequential DDES displayed a capacity for anti-thrombus, re-endothelialization, and hyperplasia inhibition.

5.3.Cell therapy stents

5.3.1.Cell seeding

Fig.8.(A-E) Cell seeded stents after expansion.Scale bars = 100 μm.(F) The illustration of MNP-laden EC which is oriented to the site of stenting in mouse.

Van Der Giessen et al.was the first to report EC seeding stents.He claimed that EC seeding stents may be a potential approach for the repair stent-related endothelium injuries [140].Conte et al.demonstrated successful EC seeding on the artery wall,confirming its function in promoting endothelium restoration at the site of acute arterial injury [141].Thus, seeding ECs onto the surface of stents and the local delivery of ECs to the artery wall is an attractive approach for the treatment of an injured endothelial layer.Wu et al.researched the potential mechanism of re-endothelialization of EC seeded stents, as shown in Fig.8A-E [142].They transfected HUVECs with VEGF and seeded them on the surface of a stent,and found that VEGF-overexpressed HUVECs promoted EC growth and inhibited hyperplasia in 1 week compared to BMS, indicating the potential of a VEGF-mediated re-endothelialization mechanism.Moreover, stents seeded with mesenchymal stem cells(MSCs) or endothelial progenitor cells (EPCs) represent another effective approach to promoting re-endothelialization after mechanical injury [143,144].Although re-endothelialization and biocompatibility have been verified in many in vitro experiments,several studies have found failure in vivo due to the quick loss of seeded cells, seeded ECs shedding under expansion, a low adherence to the artery wall, and cell retention due to blood flow.Surface modifications were developed to enhance the stability of ECs anchored to vessel walls via a biological matrix.Kipshidze et al.fabricated a fibrin glue matrix to anchor ECs, resulting in an endothelial coverage of around 70%,compared to 10%in the control sample.This fibrin/EC matrix resulted in a significant reduction of restenosis in an atherosclerotic rabbit model[145].Furthermore,Polyak et al.developed magnetic nanoparticle MNPs loaded with ECs for targeting mechanical injuries on vessel walls induced by stenting, dismissing the shortcomings of in situ EC seeded stents,as mentioned above,contributing to the enhanced efficiency of cell therapy to prevent ISR compared to non-treated stents (Fig.8F)[146].

5.3.2.Cell capture

To date,EPCs derived from circulating peripheral blood are considered a key contributor to early stage re-endothelialization.The recruitment of EPCs to injured sites promotes re-endothelization and inhibits neointimal hyperplasia post-stent implantation[147,148].GenousTMEPC capture stent (OrbusNeich, Fort Lauderdale,Florida,USA)is able to capture circulating EPCs by immobilizing CD34 antibodies onto the surface of stainless stents,targeting CD34+cells (i.e.EPCs) in the vascular circulation [148].The HEALING-FIM I & II and E-Healing clinical study verified the safety, feasibility, and efficacy of GenousTMEPC capture stents,reporting on the reduction of the formation of neointima in a 6-month follow-up period (late luminal loss (0.53 ± 0.06) for GenousTMEPC capture stent VS (1.01 ± 0.07) for low EPC titers)[149,150], and low incidences of revascularization and stent thrombosis [151].In addition, COMBO dual-therapy stents combining CD34 antibody and sirolimus was designed to capture bone marrow-derived EPCs via CD34 and suppress SMC proliferation via sirolimus.The clinical results indicated the non-inferiority of the bio-engineered COMBO stent (0.39 ± 0.45) mm in terms of late luminal loss compared with the 1G paclitaxel-eluting stent.However, COMBO was inferior with regards to target vessel failure and neointimal hyperplasia thickness compared to the 2G everolimus-eluting stent [152].Although the COMBO stent has plenty of room for improvement, the COMBO stent has shown a remarkable safety profile with a low incidence of target lesion failure and stent thrombosis, supporting the feasibility of short-term DAPT [153].Recently, nano-technology has been used to capture ECs, as reported by Nguyen et al., who developed a multitargeting urethane-doped polyester (UPE) nanoparticle scaffold system with dual ligands glycoprotein 1b (GP1b) and anti-CD34 antibody to target injured endothelium and capture EPCs [154].400-nm nanoparticles were an effective target for von Willebrand factor-coated surfaces featuring injured vascular wall surface, as shown in Fig.9A and B.Via the synergistic effects of the two ligands, over 57 % of neointimal hyperplasia were inhibited and 60%of re-endothelialization within 60 d was achieved.In addition to the CD34 antibody, a recent study found that the CD133 antibody results in greater EPC recruitment and adhesion [155].To promote late stage re-endothelialization, Park et al.designed an advanced EPC capture stent covered with anti-CD146 antibody loaded with silicone nanofilament for the highly efficient and specific capture of late stage EPCs,as shown in Fig.9C[156].Stent loaded with CD146 displayed markedly higher rates of late EPC capture, as well as MSCs, leading to enhanced reendothelialization and protection against ISR (Fig.9D).Moreover,the REDV peptide, a sequence of fibronectin that specifically recruits EPCs by binding to the αvβ3 integrin receptor on EPCs,exhibited high levels of EPC capture and promoted EC proliferation.These findings indicate another promising strategy for the capture of EPCs [77,157].

6.In situ Gene-Eluting Stents

Fig.9.(A)Fluorescent images of arteries with lumen injury after incubating with control particles and MTNs.(B)The fluorescent intensity was determined and compared for three groups:control(no NP),control NP(unconjugated NPs),and MTNs.(C)The illustration of silicone nanofilament(SiNf)with CD-146 Ab grafted for capture of EPC.(D)The in vivo results of porcine stent implantation at 4 weeks.

Fig.10.Gene delivery strategies toward in-stent restenosis treatment is depicted as naked transgene delivery, non-viral vector transgene delivery and viral transgene delivery.Through delivering and transfection gene in the host cell, the transgene can express therapeutic proteins that beneficially regulate vascular responses.

Genetic technologies offer a novel and encouraging way to treat cardiovascular diseases through the delivery of transgenes (DNA,RNA, and antisense oligonucleotides) to host cells by targeting transgene expression, as shown in Fig.10 [158].Compared with traditional therapeutics,gene therapy is able to upregulate or block target gene activities and inhibit transgene protein expression,which is a crucial feature among all types of cellular events[159].For in situ therapeutic gene delivery, a novel route comprised of loading transgenes on a stent platform and introducing transgenes to injured vessel walls aims to inhibit neointimal hyperplasia [160].Gene-eluting stents can transmit promoter/enhancer sequences (transgene) to injured vessel wall and target transgene expression via vectors that induce the expression of therapeutic proteins.By upregulating or downregulating proteins with deficient or morbigenous functions in the responses between the vessels and stents, the gene eluting system offer a promising clinical outlook.Compared with other routes of vascular gene delivery to inhibit ISR (intravenous, intracoronary injection, and balloon catheter delivery), in situ gene-eluting stents exhibit improved gene transfer efficiency: (1) transgenes reach injured arteries directly via stent implantation, wherein genes are able to maximize site-specific delivery; (2) in situ gene-eluting systems are strictly localized and specifically expressed,barely disseminating to the lungs, liver, or other organs, thus preventing systemic toxicity [161]; (3) the stent platform serves as a gene reservoir to protect genes from washing away by the blood flow, allowing for their long-term release.To date, gene therapy has demonstrated its ability to modulate the phenotypic properties of SMCs, as well as their proliferation and migration, in addition to their interactions with other cells,such as ECs,inflammatory cells,and platelets[162,163].This approach could be used to modulate the proliferation and migration of SMCs, control inflammation, and accelerate endothelium healing with minimal negative effects on the regeneration of the luminal endothelial layer.

6.1.Naked DNA delivery

Gene treatment was applied on the eluting stent system by imbedding the transgene directly(DNA,RNA,or antisense oligonucleotides)into the polymeric matrix,as in the drug-eluting system.Walter et al.used the naked DNA,including the segment sequence of the human vascular endothelial growth factor (phVEGF-2), to repair the injured artery based on the PC polymeric matrix.The tailored gene segment successfully enhanced the expression of VEGF in ECs and exhibited an efficient re-endothelialization healing process [164].TGF?1, FasL, eNOS, iNOS, and TIMPs were investigated and verified as potential anti-ISR therapeutic transgenes with anti-proliferation, anti-inflammation, and re-endothelialization properties[165-167].Therefore,in situ gene therapy is a promising strategy for preventing ISR.However, this method is associated with a low gene transfer efficacy, rapid degradation by nucleases,and clearance by mononuclear phagocytes.

6.2.Vector DNA delivery

6.2.1.Non-viral vector

Vector are able to transverse host cells and have the capacity to carry large DNA.Therefore,vectors are used to enhance the permeability of exogenous transgenes.Currently, vectors used in gene stents are classified as either non-viral vectors or viral vectors.Among the non-viral vectors, the dominating non-viral vectors are synthetic plasmids,liposomes,and polymer complex.Plasmids are circular double stranded DNA oligonucleotides that are separate from but can replicate independently in the host cell’s chromosome.By loading transgenes, the plasmid vector has verified the significant improvement upon the uptake efficiency of transgene into the host cell cytoplasm.For instance,Egashira et al.delivered 7ND transgene-plasmid via a gene-eluting stent to the local artery.By expressing the targeted transgene, this gene-eluting stent inhibited the chemotaxis of mononuclear leukocytes and the proliferation and migration of SMCs.An animal model study also evidenced that in situ gene-eluting stents can effective attenuate ISR[168].As mentioned above,NO,a multifunctional cytokine derived from ECs, has anti-platelet, anti-SMC proliferation, antiinflammation properties [110].Notably, the lack of NO at the site of vascular injury may be a pivotal factor contributing to ISR.Ji et al.designed a gene delivery system with a plasmid DNA vector encoding short hairpin RNA (shRNA) to downregulate the TGF?1 expression which plays a key role in enhancing NO production[169].In addition,Sharif et al.utilized liposome to load eNOS genes based on stents, which showed enhanced re-endothelialization by upregulating NO generation compared to PC stents [170].

Notably,polymer complexes are a promising alternative to vectors used in eluting stent systems.They have attracted research attention in recent years due to their efficiency as passing barriers.Yang et al.used PLGA nanoparticles to load dual agents:VEGF gene to target ECs healing and paclitaxel(PTX)to inhibit SMC proliferation [171].The VEGF/PTX NP-coated stents showed complete reendothelialization and significantly suppressed ISR in vivo.Recently, Ye et al.synthesized a reduction-responsive branched nucleic acid vector (SKP) to load pVEGF transgene, constructing a SKP/pVEGF complex eluting stent, as shown in Fig.11A [172].The rabbit experiment evidenced the outstanding performances of the SKP/pVEGF stent in terms of anti-ISR properties and reendothelialization by effectively expressing VEGF protein in vivo.Moreover, responsive and smart polymeric vectors were recently reported, with the aim of treating abnormal microenvironments in the process of endothelium healing, such as matrix metalloproteinase (MMP) responsive vector [173].Upon the activation of MMPs,such as MMP2 and MMP9,a marked increase was observed in the ISR model, which has been identified to regulate cell proliferation, migration, and accelerate interaction between cells and ECM[174].However,non-viral vectors still yield an unsatisfactory transfer efficacy due to their inability to insert DNA into the cell nucleus, especially in non-diving cells, as well as the rapid degradation of vectors in vivo.

6.2.2.Viral vector

Compared with non-viral vectors, viral vectors are associated with a higher efficacy of delivery to target cells.Adenoviral vector(AV), the most widely used gene vector in gene delivery systems,carries a transgene into the cytoplasm via specific receptors(Fig.10).For gene eluting stents, adenoviral vectors encoding human inducible nitric oxide synthase (AdiNOS) can induce the overexpression of iNOS at the injured site for to enhance NO,repair ECs and reduce neointimal hyperplasia[175].The endothelial nitic oxide synthase(eNOS)has a similar effect as iNOS[176].To ensure the controlled release of AV from the stents, Fishbein et al.designed a gene vector carrier matrix(PABT/PEI(PDT))with AdiNOSto bind to bare-metal platform surfaces via a reversible ester bond(HL) [177].This reversible immobilization route allows for the constant and stable release of gene vectors to vascular vessels, as well as efficient gene expression.However,it should be noted that overexpressing iNOS in vivo tends to results in the formation of O2-, which can hinder vascular repair and increase neointima formation.

However, due to the immune response and short-term gene transfer of AV of viral genomes in vivo, the new generation viral vectors, such as the adeno-associated virus (AAVs), RNA retroviruses, and lentivirus, have been developed, with a low immune response and prolonged gene expression,maintaining the integrity of the gene in the host genome [178].However, the low yield of AAVs is an important limitation for large-scale applications.In terms of retroviruses, they are only able to transfer proliferative cells and are harder to produce than Ad.At present, nanoparticle technology, in particular magnetic particles, are attractive for the treatment of cardiovascular diseases, improving the therapeutic impacts and reducing the off-target effects.Vosen et al.used lentiviral vectors (LV) and magnetic nanoparticles (MNP) in complex combined with tailored systematic magnet to overexpress the vasoprotective gene(eNOS)in ECs(Fig.11B)[179].The MNPs were successfully observed in the cytoplasm of ECs after 72 h of transduction (eGFP expression detected).These MNP-loaded and eNOS-overexpressing ECs could be magnetically navigated to a site-specific position via magnetic fields in a radially symmetric manner under flow conditions.

Fig.11.(A) Schematic illustration of the preparation of S-SKP/pVEGF and its in vivo implantation process in rabbit.(B) Strategy of radially symmetric EC replacement in mouse aortas ex vivo using lentivirus (LV)/magnetic nanoparticle (MNP) complexes.

Despite many gene therapies having been studied for the treatment of cardiovascular diseases, the majority are limited by a low gene transfer efficiency and different levels of immune responses,leading to a low physiological effect, insufficient distribution of vectors, and insufficient gene expression, followed by fast vessel regression.Moreover,the blood flow may wash vectors away from stents.Therefore,there is still room for improvement in the development of an in situ stent for gene delivery.Table 3 summarizes vectors currently used to treat cardiovascular diseases.Modifying the vectors can result in gene vectors with a better efficiency and biocompatibility.Fishbein et al.fabricated a sustained AdiNOSeluting system via the covalent immobilization of thiolated protein G,which binds to the Fc fragment of mammalian IgG.The thiolated protein G was specifically bound to the vector-capturing antibodies and subsequent vectors [180].These results demonstrate the great potential of anti-restenosis abilities in vivo.To date, preclinical animal data have showed a promising perspective of in situ gene eluting stents for the treatment of ISR.However,human clinical trial have yet to begin.

7.Conclusions and Perspectives

Several strategies have been investigated for the treatment of ISR post-stenting, including interdisciplinary combinations.Although 1G and 2G DESs markedly inhibit neointima, the longterm safety profiles(≥1 year) are seriously hindered by new complications, such as LST, due to the toxicity of drugs and polymerinduced inflammation.This review aimed to systematically summarize currently used strategies and find a better to prevent ISR.The different strategies have made breakthroughs in different ways, introducing the new ideas for the treatment of ISR in addition to drug inhibition.Currently, establishing a pro-healing function has become a popular challenge,due to the fact that rapid reendothelialization possesses an excellent biocompatibility to suppress neointima hyperplasia and prevent late stage complications by modulating the secretion of cytokines.Novel strategies based on cell and gene therapy hold great promise for the selective targeting of cells and the regulation of cellular activity, enhancing both the safety and efficacy profiles of cardiovascular stents.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

The authors wish to acknowledge financial support from the National Key Research and Development Program of China(2017YFB0702500), Natural Science Foundation of China (NSFC Project, 81801853) and Sichuan Science and Technology Program(19GJHZ0058).